Exposure to SARS-CoV-2 and Infantile Diseases

Background and Aim  Immune response against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in newborns and children after prophylactic immunization is currently a relevant research topic. The present study analyzes the issue by examining the possibility that the anti-SARS-CoV-2 immune responses are not uniquely directed against the virus but can—via molecular mimicry and the consequent cross-reactivity—also hit human proteins involved in infantile diseases. Methods  Human proteins that—if altered—associate with infantile disorders were searched for minimal immune pentapeptide determinants shared with SARS-CoV-2 spike glycoprotein (gp). Then, the shared pentapeptides were analyzed for immunologic potential and immunologic imprinting phenomena. Results  Comparative sequence analysis shows that: (1) numerous pentapeptides (namely, 54) are common to SARS-CoV-2 spike gp and human proteins that, when altered, are linked to infantile diseases; (2) all the shared peptides have an immunologic potential since they are present in experimentally validated SARS-CoV-2 spike gp-derived epitopes; and (3) many of the shared peptides are also hosted in infectious pathogens to which children can have already been exposed, thus making immunologic imprint phenomena feasible. Conclusion  Molecular mimicry and the consequent cross-reactivity can represent the mechanism that connects exposure to SARS-CoV-2 and various pediatric diseases, with a fundamental role of the immunologic memory and the history of the child's infections in determining and specifying the immune response and the pathologic autoimmune sequela.


Introduction
Recently, researchers and clinicians called attention on the issue of vaccinating newborns and children against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) to protect from coronavirus disease 2019 . Pros and cons have been examined and discussed in light of the following data: • Children account only for 1.7 to 2% of the diagnosed cases of COVID-19. 1 • COVID-19 in children shows a milder disease course and better prognosis than adults. Mortality is extremely low. 2 • COVID-19 is deadlier for aged people than for other age groups. 3 the so-called multisystem inflammatory syndrome in children (MIS-C). MIS-C is rarest and rarely is fatal. Indeed, Ergenc et al 5 reported that among 1,340 patients aged between 0 and 216 months and diagnosed with COVID-19, only 6 patients had MIS-C, which corresponds to a MIS-C incidence of 0.4%. None of the patients died. In parallel, Payne et al 6 reported that MIS-C incidence per 1,000,000 SARS-CoV-2 infections was 316. • Moreover, and crucially, an analysis of the potential risk of autoimmune cross-reactivity 7-17 lacks while it should be compulsorily included in the benefit-risk assessment of SARS-CoV-2 vaccination in children.
Then, the present study aims at exploring (or excluding) the possibility that molecular mimicry and the consequent potential cross-reactivity may exist between SARS-CoV-2 and human proteins that are linked, when altered, to infantile diseases, so that targeting the viral antigen via vaccination might equate to hit human proteins linked to childhood pathologies.

Materials and Methods
Peptide sharing between SARS-CoV-2 spike glycoprotein (gp) (NCBI, GenBank Protein Accession Id ¼ QHD43416.1) from SARS-CoV-2 and human proteins related to childhood diseases was analyzed as previously detailed 9,10 using the pentapeptide as minimal immune determinant unit. Pentapeptides were used as sequence probes since a peptide grouping composed of five amino acid (aa) residues defines a minimal immune unit that can (1) induce highly specific antibodies and (2) determine antigen-antibody-specific interaction. 18,19 A library of 372 human proteins linked-when altered-to pediatric diseases was obtained from UniProt database (www.uniprot.org) 20 using the keyword "infantile." The 372 proteins are listed in ►Supplementary Table S1 (online only).
Methodologically, the SARS-CoV-2 spike gp primary aa sequence was dissected into pentapeptides offset each other by one residue (i.e., MFVFL, FVFLV, VFLVL, FLVLL, and so forth) and the resulting viral pentapeptides were analyzed for occurrences within the human proteins related to infantile diseases. Peptide Match and Peptide Search programs available at www.uniprot.org 20 were used.
The immunologic potential of the peptides shared between SARS-CoV-2 spike gp and proteins related to childhood diseases was analyzed by searching the Immune Epitope Database (IEDB, www.iedb.org/) 21 for immunoreactive SARS-CoV-2 spike gp-derived epitopes hosting the shared pentapeptides.

Results and Discussion
Molecular Mimicry between SARS-CoV-2 Spike gp and Human Proteins Related to Infantile Diseases ►Table 1 shows that SARS-CoV-2 spike gp shares a high number of minimal immune determinants (namely, 54) with 43 human proteins that associate with infantile disorders when altered, mutated, or, however, improperly functioning. The following points emerge from ►Table 1: • The unexpectedly high molecular mimicry described in ►Table 1 and the consequent potential cross-reactivity support the hypothesis that several diseases might occur in children following exposure to the SARS-CoV-2 antigen. • Mathematically, the vastness of the common molecular platform stands out when one considers that the probability for 2 proteins to share 1 pentapeptide on the basis of the 20 aa and neglecting the relative aa abundance is equal to 1 out of 20 raised to 5. That is, it is equal to 0.0000003125.   • Such unexpected massive peptide commonality between SARS-CoV-2 antigen gp and the human proteome indicates and confirms a strict phenetic relationship between viruses and the origin of eukaryotic cells according to the endosymbiotic theory. 63 • Pathologically, the diseasome that might occur via crossreactivity includes severe disorders such as nephropathies, seizures, cardiomyopathies, parkinsonism-dystonia, global developmental delay, hypotonia, hearing loss, ataxia, hyporeflexia, hypothyroidism, pancreatic insufficiency, and lethal encephalopathy, inter alia.

Immunologic Potential of the Peptide Sharing between SARS-CoV-2 Spike gp and Human Proteins Related to Infantile Diseases
The cross-reactivity potential of the peptide sharing described in ►Table 1 appears to be supported by inspection of IEDB (www.iedb.org). 21 Indeed, all the 54 minimal immune determinants common to SARS-CoV-2 spike gp and human proteins related to infantile diseases occur and repeatedly recur in 839 SARS-CoV-2 spike gp-derived epitopes, of which ►Table 2 displays only a synopsis in the interest of brevity. In essence, ►Table 2 validates, likely enough, the cross-reactivity hypothesis at the basis of the present study.

Specificity of the Peptide Sharing between SARS-CoV-2 Spike gp and Human Proteins Linked to Infantile Diseases
To control the specificity of the peptide sharing between SARS-CoV-2 spike gp and human proteins linked to childhood diseases (►Table 1), the 54 shared pentapeptides were analyzed for occurrences in other coronaviruses not associated with particular pediatric complications, that is, MERS-CoV, HCoV-OC43, and HCoV-229E. Results are shown in ►Table 3 that provides evidence that the intense peptide overlap between SARS-CoV-2 spike gp and human proteins related to childhood diseases is highly specific. De facto, almost all the 54 shared pentapeptides are absent in the CoV controls, that is, in the pathogenic MERS-CoV 64 as well as in the scarcely pathogenic HCoV-OC43 and HCoV-229E that cause only mild symptoms. 65

Occurrence in Bacteria of Peptides Common to SARS-CoV-2 Spike gp and Proteins Linked to Infantile Diseases
To further control the specificity of the peptide sharing between SARS-CoV-2 spike gp and human proteins linked to childhood diseases, comparative sequence analyses were extended to the bacterial pathogens B. pertussis, C. diphtheriae, C. tetani, H. influenzae, and N. meningitides-that is, bacteria to which children may be exposed also following current vaccinal routes-were analyzed. Results are displayed in ►Table 4.
It can be seen that many of the 54 peptides shared between SARS-CoV-2 spike gp and human proteins linked to infantile diseases also occur in the analyzed microbial organisms, thus highlighting that, contrary to expectations, while practically no phenetic similarity exists between SARS-CoV-2 spike gp and the control CoVs (►Table 3), a high level of similarity exists between SARS-CoV-2 spike gp and bacterial pathogens to which children have been exposed via passive/active infection and by which the immune system has already been imprinted (►Table 4). Then, as highlighted in the literature, 66-72 the interpathogen peptide commonality can add a further potential burden to the molecular mimicry phenomenon described in ►Table 1. Indeed, a fundamental property of the immune system is the memory for immune determinants previously encountered so that, as a rule, the immune system reacts by recalling memory of the responses toward past infections rather than producing ex novo responses toward the recent ones.
Consequently, in the case under study here, the following sequence of events may unfold: • A primary response to SARS-CoV-2 can actually occur as a secondary (or even tertiary) response against pathogens previously encountered and memorized by the immune system. That is, anamnestic secondary antibacterial responses can occur after exposure to SARS-CoV-2. Such anamnestic secondary antibacterial responses will be of considerable proportions given the extent of the viral versus bacterial peptide overlap described in ►Table 4.
• SARS-CoV-2 antigen will not be affected in that the immunologic memory deflects the immune response toward the already encountered peptides, that is, the bacterial peptide platform detailed in ►Table 4. • However, also the attack against the early sensitizing bacterial pathogens can fail by being the early sensitizing bacterial pathogens no more present in the organism. • Then, the ultimate result might be that the anamnestic, high affinity, high avidity, and extremely potent secondary immune response elicited by the lastly encountered pathogen-that is, SARS-CoV-2-can hit the only available targets, that is, the common immune determinants that in this instance are present in the human proteins related to infantile diseases (►Table 1).
According to this sequence of events, molecular mimicry and immunologic memory might explain also the different pathological outcomes of the autoimmune responses-from mild symptoms to even lethal pathologies-that may follow exposure to SARS-CoV-2. In practice, the history of infections/immunization of each child is the main factor dictating the onset and the severity of the pathologies outlined in ►Table 1.

Conclusion
The present study investigates the possible adverse events that might occur in newborns and children following exposure to SARS-CoV-2. Based on the extensive peptide sharing between SARS-CoV-2 gp antigen and human proteins related to infantile diseases, supported by epitopic data that confer a high immunoreactivity to the peptide sharing, and given the possibility of immunologic imprinting phenomena, this study leads to predict that exposing newborns and children to SARS-CoV-2 might associate with infantile severe diseases such as growth retardation, abnormal genitalia, epilepsy, seizures, cardiomyopathies, hypotonia, visual loss, hypercalcemia, ataxia, infantile parkinsonismdystonia, below average general intellectual functioning, encephalopathies, and inter alia. Then, the present data suggest that extreme caution be exercised in planning and implementing a mass anti-SARS-CoV-2 vaccination of infants and children.

Funding
None.